The mid-band in polyacrylymide gel electrophoresis, which showed abnormal band between the bands of LDL and VLDL, is interpreted as an increase in IDL and an increase in cholesterol content of the VLDL fraction. The determine the relationship between mid-band appearance and coronary artery disease, we measured coronary artery stenosis, mid-band, several metabolic variables, and the distribution of abdominal fat by CT scans of 161 male patients who had undergone coronary angiography. The prevalence of mid-band in the group with coronary artery stenosis was significantly higher than in the group without coronary artery stenosis. Midband appearance was associated with triglyceride concentration, insulin concentration, and inversely HDL-cholesterol concentration. Mid-band prevalence was higher when visceral fat accumulation and hypertension were found in combination. Within the hypertensive group, the cardscore was significantly higher for combined visceral fat accumulation and mid-band appearance, while in the normotensive group mid-band appearance had no effect on the cardscore. With respect to metabolic profiles in the hypertensive group, plasma insulin levels and the glucose levels were elevated in the positive mid-band and visceral fat accumulation groups.
We concluded that mid-band appearance was not atherogenic for men. It is not atherogenic when the abnormality is isolated, that is, existing in nonvisceral fat accumulation and normotension, and it is the most atherogenic when the positive mid-band, visceral fat accumulation and hypertension are all present. Also insulin resistance and/or hyerinsulinemia played a central role in the atherogenecity in the group with positive mid-band, visceral fat accumulation and hypertension.

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We studied the lipid transfer protein (LTP) activity in 23 men with old myocardial infarction (OMI) and 25 outpatients without ischemic heart disease as the control. LTP activity was measured according to Albers' method. Lipoproteins were fractionated by ultracentrifugation, and the lipids were measured by an enzymatic method. Apolipoprotein levels were measured by the TIA method. Compared with control plasma, OMI plasma showed significant decreases in HDL-C (p=0.0003) and Apo-A1 (p=0.0002) and significant increases in TG (p=0.0070), VLDL-C (p=0.0022) and Apo-B (p=0.0070). There was a significant increase in LTP activity in OMI plasma (p=0.0157). LTP activity was negatively correlated with HDL-C (r=-0.3384, p=0.0247), Apo-A1 (r=-0.3969, p=0.0084) and the HDL-C/HDL-TG ratio (r=-0.4352, p=0.0025). From these results we concluded that in patients with OMI, high LTP activity may lead to decreased plasma HDL-C and changes in HDL composition.

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It has been reported that several DNA mutations in the receptor binding domain of apolipoprotein E (apo E) are usually involved in type III hyperlipoproteinemia. GC-clamp denaturing gradient gel electrophoresis (DGGE) and single-strand conformation polymorphism (SSCP) were used to detect DNA sequence changes. We evaluated these methods ability to determine apo E isoforms or detection of new apo E mutation. Genomic DNA for polymerase chain reaction (PCR) was extracted from whole blood of 200μl. A 244-bp fragment containing amino acid residues from 91 to 165 was amplified by first PCR. This DNA fragment included the receptor binding domain. It also included 112 and 158 amino acid residues whose mutations characterize the two common variants, apo E2 and apo E4, respectively. The second PCR product was amplified with 5'-primer or 3'-primer, which had been attached with a 40-bp G+C-rich sequence (GC-clamp), and the product was subjected to electrophoresis using a denaturing gradient gel in a bath at 60°C. The gel consisted of 7% polycrylamide with a linearly increasing gradient from 60% to 80% denaturant (100% denaturant: 7M urea/40% formamide) for 5'-GC-clamp or from 50% to 70% denaturant for 3'-GC-clamp. After electrophoresis, the gel was stained with ethidium bromide. The 5'-GC-clamp-DGGE made it possible to distinguish E4 and E3 from E2. The 3'-GC-clamp-DGGE distinguished E3 from E4. All six phenotypes of apo E could be determined by combining the 5'-and 3'-GC-clamp-DGGEs. Additionally, apo E-Kochi (145Arg->His), a rare mutation of apo E, could be detected using these methods.
SSCP was performed nonradioactively. The first PCR product was denatured to single strands by 0.5 M NaOH and heating at 42°C, and was resolved with 100% formamide. The sample was subjected to electrophoresis using 6% polyacrylamide gel at 4°C for 80 minutes. The gel was then stained using ethidium bromide. The SSCP of apo E2 homozygote showed one band, those of E3 and E4 homozygote showed two bands. The SSCP of heterozygotes showed three to four bands, whose patterns all differed. Therefore, the six phenotypes of apo E were clearly distinguished by this method. In the SSCP method, the DNA of apo E-Kochi showed yet another pattern. The SSCP method for apo E analysis was simpler and more sensitive than DGGE. PCR-SSCP took only 6 hours to analyze. Fifty patients with hyperlipoproteinemia were studied using these two methods, and another family of apo E-Kochi was found. It shoud be possible to find other mutations of apo E using these methods.

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The effects of obesity and diabetes mellitus on blood pressure have not been fully elucidated. We studied the influence of the body mass index, glucose intolerance, age, and sex on the prevalence of hypertension in 5546 sujects who had taken health examinations.
The subjects were divided into 5 groups according to the body mass index : thin, somewhat thin, somewhat overweight, overweight, and obese. They were also divided into 4 groups using an oral glucose tolerance test: normal (NOR), borderline (BOR), impaired glucose tolerance (IGT), and diabetes mellitus (DM). Thereafter we checked the hypertensive cases of each subgroup in accordance with WHO criteria.
The following results were obtained:
1) Men 40 to 49 years old
a) No hypertensive cases were shown in the thin subgroup with NOR.
b) The prevalence of hypertension in the somewhat overweight subgroup with NOR was significantly higher than in the thin and somewhat thin subgroup with NOR.
3) The prevalences of hypertension in the somewhat thin subgroup with IGT and DM was significantly higher than in this subgroup with NOR and BOR.
2) Men 50 to 59 years old
a) The prevalences of hypertension in the overweight and obese subgroup with NOR was significantly higher than in the thin or somewhat thin subgroup with NOR.
b) The prevalence of hypertension in the somewhat overweight subgroup with DM was significantly higher than in this subgroup with NOR.
3) Men 60 to 69 years old
a) The prevalence of hypertension in the obese subgroup with NOR was significantly higher than in the thin and somewhat thin subgroup with NOR.
b) The prevalence of hypertension in the somewhat overweight subgroup with DM was significantly higher than in this subgroup with NOR.
4) Female
a) No significant findings were made between the 5 subgroups by body mass index level with NOR. We concluded the following:
(i) The prevalence of hypertension was high even in the early stages of obesity.
(ii) Diabetic cases were frequently accompanied by hypertension even without obesity.

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It has been accepted that the incidence of coronary artery disease (CAD) in patients with hypercholesterolemia (HC) can be lowered by reducing serum cholesterol levels. However, this concept is based mainly on the studies designed in the United States and Europian countries. There are few reports about the effects of medical treatment for HC in Japan.
The purpose of our study was to clarify the effects of and problems associated with medical treatment for HC in the Department of Geriatrics, Keio University Hospital.
We enrolled 230 patients treated medically with probucol, pravastatin, and/or other drugs including clofibrate, colestyramine. We excluded patients who had taken drugs for hypertension, diabetes mellitus, or CAD. Patients with undiagnosed chest pain and treated medically within 24 months were also excluded. Finally, 80 asymptomatic patients treated medically over 24 months were monitored for another 24 months. The endpoint of this study was the occurrence of CAD, cancer, or other cardiovascular disease.
The serum cholesterol level before treatment was 301±6 mg/d/ and it decreased to 242±5 mg/dl (-19%). LDL-cholesterol also decreased by 23%, but HDL-cholesterol didn't change with treatment. CAD developed in 6 patients during a 2-year follow-up period. There was no difference in total cholesterol, LDL-cholesterol, and triglyceride levels between patients with or without the development of CAD before treatment, at the start the of trial, and at the end of the trial; however, HDL-cholesterol levels at the start and end of the trial were significantly lower in patients with newly developed CAD. Therefore, 4 of 6 newly developed CAD patients were associated with an excessive decrease in HDL-cholesterol.
The sample size of our study was not large enough to draw definitive conclusions, but the results suggested that medical treatment with probucol, pravastatin, or other drugs was effective in lowering serum cholesterol and LDL-cholesterol levels. Our study also suggested that possible problems may be associated with the treatment of HC. Excessive decreases in HDL-cholesterol level during treatment could lead to the development of CAD, regardless of the choice of drug. The normalization of both total cholesterol and HDL-cholesterol levels could be equally important in treatment for HC.

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Raised lipoprotein (a) [Lp(a)] concentrations have been reported in patients with non-insulin dependent diabetes mellitus (NIDDM), especially those with diabetic nephropathy. The purpose of this study was to demonstrate a direct relationship between Lp(a) levels and arteriosclerosis in diabetics. The relationship between plasma Lp(a) levels and aortic pulse wave velocity (PWV), glycemic control and proteinuria were studied in 133 NIDDM patients (71 men and 62 women). The incidence of a high-Lp(a) group (≥32 mg/dl) increased with the degree of proteinuria. In macroalbuminuria group, significant correlations were found between Lp(a) and PWV (r=0.55, p<0.05) and fibrinogen concentrations (r=0.53, p<0.05). Furthermore, PWV and plasma levels of fibrinogen in the high-Lp(a) group were significantly higher than those in the normal-Lp(a) group. On the other hand, in all subjects, Lp(a) levels were correlated with plasma levels of glycosylated hemoglobin A₁c, fibrinogen and the duration of the disease. When compared with the degree of glycemic control, Lp(a) levels were significantly higher in diabetics with poor glycemic control.
From these results, it seems that plasma Lp(a) levels in diabetics are affected by hyperglycemia. However, in patients with advanced diabetic nephropathy, it may accelerate the development of arteriosclerosis in association with hyperfibrinogenemia.

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We studied the occurrence of myocardial ischemia due to peripheral disturbances of coronary circulation using histological analyses and measurements of the cardiac function.
Measurements were taken for EDP, max dP/dt, max(—)dP/dt, blood pressure, pulse rate and left ventricular pressure in 10 rabbits before and after administration of a cholesterol diet for 3 months (chol rabbits), The same measurements were also taken in 5 normal rabbits. Counting was made of all the peripheral coronary arteries in diameter 50∼150 μm in which more than 75% of stenosis was observed by sectioning longitudinally every 5 mm and fibrosis % was also found.
In chol rabbits, EDP exhibited significant increases from 2.1±1.2 to 7.3±4.0 mmHg, and max(—)dP/dt exhibited significant decreases from 3, 955±583 mmHg to 3, 300±429 mmHg before and after the administration, respectively (p<0.05). No significant difference was observed in the other items, however. The main coronary artery exhibited stenosis in not more than 50% of either group, but in the chol rabbits, significant increases were observed both in the ratio of the number fo stenotic blood vessels (20.7%, 0.94%) and the fibrosis (2.71%, 1.66%).
In the chol rabbits, the increases in EDP and the decrease in max(—)dP/dt was pressumed to have resulted from peripheral coronary stenosis and associated fibrosis, and was considered to suggest the presence of myocardial ischemia.

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To evaluate the effect of EPA on arteriosclerosis, 24 Japanese white rabbits were studied as cuffsheathed models, which produced diffuse intimal thickening of the carotid artery. They were divided into 4 groups. Groups I and II were fed a normal diet, while groups III and IV were fed with a 0.5% cholesterol diet after cuff treatment. In groups I and III, EPA was administered p.o. forcibly at 600 mg/kg/day 1 week before cuff treatment. The cuff was placed around the carotid artery for three weeks. Then, carotid artery samples were obtained for pathological studies. From the elastica-stained samples, the area of the intima (AI), media (AM), and the part surrounded by the internal elastic lamella (A-IEL) were measured using a digitizer, to calculate AI/A-TEL (IT) and AI/AM (IT-M) as indices of intimal thickening.
The mean IT-M in group I (0.14) decreased significantly compared with group II (0.22), which showed cellulofibrous intimal thickening with a partly lacerated IEL. The mean IT in group III (0.30) was also significantly lower compared with group IV (0.46), which revealed prominent intimal thickening with spindleshaped smooth muscle cells. These findings suggested that EPA suppressed intimal thickening, with cell proliferation observed in the early phase of atherosclerosis.

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